Clinical next generation sequencing in developmental and epileptic encephalopathies: Diagnostic relevance of data re-analysis and variants re-interpretation.

Developmental and epileptic encephalopathies (DEE) are complex pediatric epilepsies, in which heterogeneous pathogenic factors play an important role. Next-generation-sequencing based tools have shown excellent effectiveness. The constant increase in the number of new genotype-phenotype associations suggests the periodic need for re-interpretation and re-analysis of genetic studies without positive results. In this study, we report the diagnostic utility of targeted gene panel sequencing and whole exome sequencing in 55 Argentine subjects with DEE, focusing on the utility of re-interpretation and re-analysis of undetermined and negative genetic diagnoses. The new information in biomedical literature and databases was used for the re-interpretation. For re-analysis, sequencing data processing was repeated using updated bioinformatics tools. Initially, pathogenic variants were detected in 21 subjects (38%). After an average time of 29 months, 25% of the subjects without a genetic diagnosis were re-categorized as diagnosed. Finally, the overall diagnostic yield increased to 53% (29 subjects). In consequence of the re-interpretation and re-analysis, we identified novel variants in the genes: CHD2, COL4A1, FOXG1, GABRA1, GRIN2B, HNRNPU, KCNQ2, MECP2, PCDH19, SCN1A, SCN2A, SCN8A, SLC6A1, STXBP1 and WWOX. Our results expand the diagnostic yield of this subgroup of infantile and childhood seizures and demonstrate the importance of re-evaluation of genetic tests in subjects without an identified causative etiology.

Molecular Diagnosis in an Argentinean Mitochondrial Disorders Cohort

Abstract Mitochondrial diseases are multisystemic disorders characterized by an impairment of the mitochondrial respiratory chain. Diagnosis requires an approach that involves a high index of suspicion, molecular techniques and a careful selection of the tissue to be studied. Our goal was to develop and implement local strategies for diagnosing mitochondrial disorders, by standardizing procedures of molecular biology and nucleic acid sequencing. A prospective, analytical, observational study was conducted in a cohort of, a total of 82 patients with suspected mitochondrial disorder who were treated at our hospital between May 2008 and June 2019. We developed molecular diagnostic tools that included classical monogenic techniques and Next Generation Sequencing. We characterized the neurological and extra neurological manifestations noted in our cohort. Following the proposed algorithm, we obtained a molecular diagnostic performance of 54%, identifying mutations in 44 patients. mtDNA mutations were identified in 34 patients. Structural rearrangements in mitochondrial genome were found in 3 and 7 in nuclear genes, respectively. Our results confirm the utility of the proposed algorithm and the molecular tools used, as evidenced by a high diagnostic performance. This is of great value to a more efficient and comprehensive medical care of patients and families affected by mitochondrial disorders.

SLC6A4 gene variants and temporal lobe epilepsy susceptibility: a meta-analysis.

There seems to be a role for serotoninergic neuro-transmission in the pathophysiology of the epilepsies. Different groups have studied the role of regulatory variants in the SLC6A4 gene, which code for the central serotonin transporter, in the complex genetics of temporal lobe epilepsy (TLE) obtaining contradictory findings. Therefore, a systematic review and critical analysis of this topic seem to be timely. Published studies up to October 2011 of TLE and the SLC6A4 promoter and intron 2 variant number repeat polymorphisms (VNTR) were identified by searches of Medline, Scopus and ISI-Web of Sciences databases. Meta-analysis of TLE case-control data were performed to assess the association of SLC6A4 VNTRs with TLE susceptibility. Pooled odds ratios were estimated by means of a genetic-model-free approach. The quality of the included studies was assessed by a score. The studies included compared a total of 991 TLE cases and 1,202 controls. We did not find synthetic evidence of association between SLC6A4 promoter and intron 2 variants and the risk of TLE. However, the intron 2 VNTR seems to have opposite effects in different populations. In this meta-analysis our findings were inconclusive in order to associate any of the 5-HT receptor gene variants with the risk of TLE.

ApoE epsilon4 genotype and the age at onset of temporal lobe epilepsy: a case-control study and meta-analysis.

In order to investigate the role of ApoE epsilon4 as a modifier of the age at onset of temporal lobe epilepsy (TLE), we performed a molecular epidemiology study in 78 patients with mesial temporal lobe epilepsy and hippocampal sclerosis. Genotyping was done by a PCR-RFLP assay. In order to better estimate the role of this variant as a modifier of the age at onset, we also performed a systematic review of the literature. We included our results into a meta-analysis along with data available from seven published studies with 728 patients that looked into the role of ApoE epsilon4 in TLE. We found that ApoE epsilon4 carriers in our population had a non-significant earlier age of epilepsy onset than non-carriers. The meta-analysis confirmed this finding, showing that ApoE epsilon4 carriers had epilepsy onset almost 4 years earlier than non-carriers (mean difference 5.15 years; CI 95% 2.08-6.22; p=0.001). In conclusion, the ApoE epsilon4 isoform is a genetic factor that might influence the age at onset of TLE.

Association study between interleukin 1 beta gene and epileptic disorders: a HuGe review and meta-analysis.

Previous studies have examined the association of a single nucleotide polymorphism at the promoter region of interleukin 1B (IL-1 beta-511T) with temporal lobe epilepsy and febrile seizures susceptibility, but those studies have been inconclusive. Published studies up to March 2007 of temporal lobe epilepsy, febrile seizures and the IL-1 beta-511T single nucleotide polymorphism were identified by searches of Medline and Embase databases. Meta-analysis of temporal lobe epilepsy and febrile seizures case-control data were performed to assess the association of IL-1 beta-511T with temporal lobe epilepsy, temporal lobe epilepsy with hippocampal sclerosis, febrile seizures, and other epileptic disorders. Pooled odds ratios (OR) were estimated by means of a genetic-model-free approach. The quality of the included studies was assessed by a score. The results show a modest association (OR, 1.48; 95% confidence interval, 1.09-2.00; P = 0.01) between the IL-1 beta-511T polymorphism and temporal lobe epilepsy with hippocampal sclerosis.

Is tumor necrosis factor-376A promoter polymorphism associated with susceptibility to multiple sclerosis?

A single nucleotide polymorphism (SNP) at position -376 of the tumor necrosis factor alpha gene (TNFA) has been associated with susceptibility to multiple sclerosis (MS) in Spain. However, no association was found in populations from the USA and The Netherlands. Here we investigate the association between the TNFA-376A SNP and MS susceptibility in Argentinean patients with MS. The A/G genotype was found in 4.4% of patients (n=90) and in 4.8% of healthy individuals (n=84; p=0.92; odds ratio=0.93; confidence interval: 0.23-3.84). Thus, no significant differences in genotype and allele frequencies were found between healthy individuals and patients with MS in Argentina.